Insulin-dependent diabetes mellitus (IDDM) is a leading cause of morbidity and mortality. Using an animal model of IDDM, the non-obese diabetic (NOD) mouse, we will examine the basic autoimmune process underlying IDDM and the mode of action of antigen-based immunotherapeutics. Specifically; 1). Little is known about the specificity and phenotype of the T cells that infiltrate the pancreas during the development of IDDM. We will determine when, how many and what type of antigen-specific T cell infiltrate the pancreas of NOD mice, and whether splenic autoantigen- specific T cell populations reflect those in the target tissue. 2). There is a lack of immunological markers with which to evaluate the status of B-cell autoimmunity. In particular, the relationship between PBMC and pancreatic T cell responses following immunotherapy is an open question. In the NOD mouse, we will test whether changes in the frequency of splenic (i.e., PBMC) antigen-specific Th1/Th2 cell populations, as well as changes in autoantibody isotypes following immunotherapy reflect changes in the infiltrating antigen-specific T cell population in the pancreas and can serve as predictive markers of disease outcome. 3). We have observed that the induction of Th2 responses to a single GAD determinant leads to the spreading of Th2 autoimmunity. Determinant spreading of Th2 responses may be a fundamental mechanism underlying antigen-based immunotherapeutics, explaining why different auto-antigens can be successfully used for immune therapy, as well as observations of "infectious tolerance". We will further characterize Th2 determinant spreading following antigen-based immunotherapy. 4). Characterize the mechanism(s) underlying Th2 dominant spreading. We will investigate the role of the following factors on Th2 spreading; 1) the size of the first wave of autoreactive Th2 cells, 2) the cytokine environment, and 3) the size of the antigen-specific T cell precursor pool. We will also determine whether naive, or primed and uncommitted T cells are required for Th2 spreading. Together, these studies will address basic issues concerning Th1/Th2 cell involvement in disease pathogenesis and the establishment of tolerance. The results have potential clinical utility through improving our ability to monitor and inhibit the autoimmune process underlying human IDDM.